Phosphatidylcholine transfer protein (PC-TP) is a steroidogenic acute regulatory-related lipid transfer (START) domain protein, which in vitro catalyzes the intermembrane transfer of phosphatidylcholines, exclusively. During the previous project period, we demonstrated important roles for PC-TP in biliary lipid secretion, hepatic cholesterol homeostasis and high density lipoprotein (HDL) metabolism. We also solved the three-dimensional structure of the protein complexing a phosphatidylcholine and identified the membrane interaction domain. In contrast to our original prediction that PC-TP acts to shuttle phosphatidylcholines from the endoplasmic reticulum to the canalicular plasma membrane for secretion into bile, this research has suggested a more global regulatory role in hepatic lipid homeostasis. The current proposal tests the hypothesis that PC-TP senses the fatty acyl chain composition of membrane phosphatidylcholines and engages in protein-protein interactions that control the metabolism of lipids within the liver. Preliminary studies have demonstrated that triglycerides accumulate in livers of Pctp~'~ mice and that PC-TP is highly regulated by peroxisomal proliferator-activated receptor alpha (PPARa). Specific Aim 1 will use Pctp~'~ and wild type littermate control mice to examine the influence of PC-TP on hepatic triglyceride metabolism, very low density lipoprotein (VLDL) production and the expression of hepatic genes that regulate lipid homeostasis. Mice will be challenged with a diet enriched in saturated fat,which increases hepatic production and export of triglycerides, and a diet supplemented with the PPARct ligand fenofibrate, which promotes fatty acid utilization by the liver. Specific Aim 2 will characterize the interactions between PC-TP and proteins identified in mouse liver and embryo by yeast two-hybrid screening. Specific Aim 3 will elucidate the conformational changes of PC-TP that are required to bind phosphatidylcholines, as well as the structural basis for interactions between PC-TP and other proteins. These studies should provide fundamental new insights into the biology of START domain proteins and the molecular regulation of hepatic lipid metabolism, which may potentially lead to novel strategies for the diagnosis and management of common disorders, such as nonalcoholic fatty liver disease.